Tags

Type your tag names separated by a space and hit enter

Molecular analysis of several in-house rRT-PCR protocols for SARS-CoV-2 detection in the context of genetic variability of the virus in Colombia.
Infect Genet Evol. 2020 10; 84:104390.IG

Abstract

The COVID-19 pandemic caused by SARS-CoV-2 is a public health problem unprecedented in the recent history of humanity. Different in-house real-time RT-PCR (rRT-PCR) methods for SARS-CoV-2 diagnosis and the appearance of genomes with mutations in primer regions have been reported. Hence, whole-genome data from locally-circulating SARS-CoV-2 strains contribute to the knowledge of its global variability and the development and fine tuning of diagnostic protocols. To describe the genetic variability of Colombian SARS-CoV-2 genomes in hybridization regions of oligonucleotides of the main in-house methods for SARS-CoV-2 detection, RNA samples with confirmed SARS-CoV-2 molecular diagnosis were processed through next-generation sequencing. Primers/probes sequences from 13 target regions for SARS-CoV-2 detection suggested by 7 institutions and consolidated by WHO during the early stage of the pandemic were aligned with Muscle tool to assess the genetic variability potentially affecting their performance. Finally, the corresponding codon positions at the 3' end of each primer, the open reading frame inspection was identified for each gene/protein product. Complete SARS-CoV-2 genomes were obtained from 30 COVID-19 cases, representative of the current epidemiology in the country. Mismatches between at least one Colombian sequence and five oligonucleotides targeting the RdRP and N genes were observed. The 3' end of 4 primers aligned to the third codon position, showed high risk of nucleotide substitution and potential mismatches at this critical position. Genetic variability was detected in Colombian SARS-CoV-2 sequences in some of the primer/probe regions for in-house rRT-PCR diagnostic tests available at WHO COVID-19 technical guidelines; its impact on the performance and rates of false-negative results should be experimentally evaluated. The genomic surveillance of SARS-CoV-2 is highly recommended for the early identification of mutations in critical regions and to issue recommendations on specific diagnostic tests to ensure the coverage of locally-circulating genetic variants.

Authors+Show Affiliations

Unidad de Secuenciación y Genómica, Dirección de Investigación en Salud Pública, Instituto Nacional de Salud, Bogotá 111321, Colombia; Grupo de Salud Materna y Perinatal, Dirección de Investigación en Salud Pública, Instituto Nacional de Salud, Bogotá 111321, Colombia. Electronic address: daalvarezd@unal.edu.co.Unidad de Secuenciación y Genómica, Dirección de Investigación en Salud Pública, Instituto Nacional de Salud, Bogotá 111321, Colombia; Grupo de Parasitología, Dirección de Investigación en Salud Pública, Instituto Nacional de Salud, Bogotá 111321, Colombia. Electronic address: cefrancom@unal.edu.co.Unidad de Secuenciación y Genómica, Dirección de Investigación en Salud Pública, Instituto Nacional de Salud, Bogotá 111321, Colombia. Electronic address: kdlaitond@unal.edu.co.Unidad de Secuenciación y Genómica, Dirección de Investigación en Salud Pública, Instituto Nacional de Salud, Bogotá 111321, Colombia; Centro de Investigación en Salud para el Trópico-CIST, Universidad Cooperativa de Colombia, Santa Marta 470003, Colombia. Electronic address: jose.usmec@ucc.edu.co.Programa Ciencias de la Biodiversidad, Instituto de Investigación de Recursos Biológicos Alexander von Humboldt, Bogotá 111311, Colombia. Electronic address: nfranco@humboldt.org.co.Dirección de Redes en Salud Pública, Instituto Nacional de Salud, Bogotá 111321, Colombia. Electronic address: aflorez@ins.gov.co.Grupo de Virología, Dirección de Redes en Salud Pública, Instituto Nacional de Salud, Bogotá 111321, Colombia. Electronic address: sgomezr@ins.gov.co.Grupo de Virología, Dirección de Redes en Salud Pública, Instituto Nacional de Salud, Bogotá 111321, Colombia. Electronic address: lrodriguez@ins.gov.co.Grupo de Virología, Dirección de Redes en Salud Pública, Instituto Nacional de Salud, Bogotá 111321, Colombia. Electronic address: jbarbosa@ins.gov.co.Grupo de Virología, Dirección de Redes en Salud Pública, Instituto Nacional de Salud, Bogotá 111321, Colombia. Electronic address: eospitia@ins.gov.co.Dirección de Vigilancia en Salud Pública, Instituto Nacional de Salud, Bogotá 111321, Colombia. Electronic address: dwalteros@ins.gov.co.Dirección General, Instituto Nacional de Salud, Bogotá 111321, Colombia. Electronic address: mospina@ins.gov.co.Unidad de Secuenciación y Genómica, Dirección de Investigación en Salud Pública, Instituto Nacional de Salud, Bogotá 111321, Colombia; Grupo de Salud Materna y Perinatal, Dirección de Investigación en Salud Pública, Instituto Nacional de Salud, Bogotá 111321, Colombia; Dirección de Investigación en Salud Pública, Instituto Nacional de Salud, Bogotá 111321, Colombia. Electronic address: mmercado@ins.gov.co.

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

32505692

Citation

Álvarez-Díaz, Diego A., et al. "Molecular Analysis of Several In-house rRT-PCR Protocols for SARS-CoV-2 Detection in the Context of Genetic Variability of the Virus in Colombia." Infection, Genetics and Evolution : Journal of Molecular Epidemiology and Evolutionary Genetics in Infectious Diseases, vol. 84, 2020, p. 104390.
Álvarez-Díaz DA, Franco-Muñoz C, Laiton-Donato K, et al. Molecular analysis of several in-house rRT-PCR protocols for SARS-CoV-2 detection in the context of genetic variability of the virus in Colombia. Infect Genet Evol. 2020;84:104390.
Álvarez-Díaz, D. A., Franco-Muñoz, C., Laiton-Donato, K., Usme-Ciro, J. A., Franco-Sierra, N. D., Flórez-Sánchez, A. C., Gómez-Rangel, S., Rodríguez-Calderon, L. D., Barbosa-Ramirez, J., Ospitia-Baez, E., Walteros, D. M., Ospina-Martinez, M. L., & Mercado-Reyes, M. (2020). Molecular analysis of several in-house rRT-PCR protocols for SARS-CoV-2 detection in the context of genetic variability of the virus in Colombia. Infection, Genetics and Evolution : Journal of Molecular Epidemiology and Evolutionary Genetics in Infectious Diseases, 84, 104390. https://doi.org/10.1016/j.meegid.2020.104390
Álvarez-Díaz DA, et al. Molecular Analysis of Several In-house rRT-PCR Protocols for SARS-CoV-2 Detection in the Context of Genetic Variability of the Virus in Colombia. Infect Genet Evol. 2020;84:104390. PubMed PMID: 32505692.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Molecular analysis of several in-house rRT-PCR protocols for SARS-CoV-2 detection in the context of genetic variability of the virus in Colombia. AU - Álvarez-Díaz,Diego A, AU - Franco-Muñoz,Carlos, AU - Laiton-Donato,Katherine, AU - Usme-Ciro,José A, AU - Franco-Sierra,Nicolás D, AU - Flórez-Sánchez,Astrid C, AU - Gómez-Rangel,Sergio, AU - Rodríguez-Calderon,Luz D, AU - Barbosa-Ramirez,Juliana, AU - Ospitia-Baez,Erika, AU - Walteros,Diana M, AU - Ospina-Martinez,Martha L, AU - Mercado-Reyes,Marcela, Y1 - 2020/06/04/ PY - 2020/05/27/received PY - 2020/05/31/accepted PY - 2020/6/9/pubmed PY - 2020/10/3/medline PY - 2020/6/8/entrez KW - COVID-19 diagnostic testing KW - Genetic diversity KW - Next-generation sequencing KW - RT-PCR KW - SARS-CoV-2 SP - 104390 EP - 104390 JF - Infection, genetics and evolution : journal of molecular epidemiology and evolutionary genetics in infectious diseases JO - Infect Genet Evol VL - 84 N2 - The COVID-19 pandemic caused by SARS-CoV-2 is a public health problem unprecedented in the recent history of humanity. Different in-house real-time RT-PCR (rRT-PCR) methods for SARS-CoV-2 diagnosis and the appearance of genomes with mutations in primer regions have been reported. Hence, whole-genome data from locally-circulating SARS-CoV-2 strains contribute to the knowledge of its global variability and the development and fine tuning of diagnostic protocols. To describe the genetic variability of Colombian SARS-CoV-2 genomes in hybridization regions of oligonucleotides of the main in-house methods for SARS-CoV-2 detection, RNA samples with confirmed SARS-CoV-2 molecular diagnosis were processed through next-generation sequencing. Primers/probes sequences from 13 target regions for SARS-CoV-2 detection suggested by 7 institutions and consolidated by WHO during the early stage of the pandemic were aligned with Muscle tool to assess the genetic variability potentially affecting their performance. Finally, the corresponding codon positions at the 3' end of each primer, the open reading frame inspection was identified for each gene/protein product. Complete SARS-CoV-2 genomes were obtained from 30 COVID-19 cases, representative of the current epidemiology in the country. Mismatches between at least one Colombian sequence and five oligonucleotides targeting the RdRP and N genes were observed. The 3' end of 4 primers aligned to the third codon position, showed high risk of nucleotide substitution and potential mismatches at this critical position. Genetic variability was detected in Colombian SARS-CoV-2 sequences in some of the primer/probe regions for in-house rRT-PCR diagnostic tests available at WHO COVID-19 technical guidelines; its impact on the performance and rates of false-negative results should be experimentally evaluated. The genomic surveillance of SARS-CoV-2 is highly recommended for the early identification of mutations in critical regions and to issue recommendations on specific diagnostic tests to ensure the coverage of locally-circulating genetic variants. SN - 1567-7257 UR - https://www.unboundmedicine.com/medline/citation/32505692/Molecular_analysis_of_several_in_house_rRT_PCR_protocols_for_SARS_CoV_2_detection_in_the_context_of_genetic_variability_of_the_virus_in_Colombia_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S1567-1348(20)30221-5 DB - PRIME DP - Unbound Medicine ER -